The free base of trimetrexate, chemically named 2,4-diamino-5-methyl-6-[(3,4,5-trimethoxyanilino)methyl] quinazoline, has the following structure: ##STR1##
Trimetrexate is an inhibitor of dihydrofolate reductase (DHFR), an enzyme which catalyzes the reduction of intracellular dihydrofolate to the active coenzyme tetrahydrofolate. Inhibition of DHFR results in the depletion of this coenzyme, and thus inhibits folate-dependent formyltransferases and interferes with thymidylate and purine biosynthesis. The end result is disruption of DNA, RNA, and protein synthesis, with consequent cell death.
Trimetrexate free base decomposes over time when exposed to oxygen at room temperature. The complete mechanism by which this process occurs is not known, although the formation of 6-aminomethyl-5-methyl-2,4-quinazolinediamine has been observed during trimetrexate degradation. Interestingly, it has been reported that a major decomposition product of the trimetrexate glucuronate salt in solution is (2,4-diamino-5-methyl-6-carboxaldehyde)quinazoline. Stetson, P. L., et al., J. Chromatography, 464, 163-171 (1989). Formation of both of these decomposition products is catalyzed by heat and light. Cf March, J., Advanced Organic Chemistry pp. 1194-1195 (4.sup.th ed., 1992).
An injectable form of the amorphous glucuronate salt of trimetrexate is sold under the commercial name Neutrexin.RTM. by U.S. Bioscience (West Conshohocken, Pa.). Neutrexin.RTM. provides trimetrexate glucuronate as a lyophilized powder suitable for reconstitution, which, when reconstituted, is stable for 48 hours at room temperature, 4 days when refrigerated, or 8 days when stored at freezer temperatures. Physicians' Desk Reference, 53rd ed., pp. 3172-3175 (1999).
Neutrexin.RTM. has been approved for use in the United States and Canada for the treatment of Pneumocystis carinii pneumonia (PCP) in patients with acquired immune deficiency syndrome (AIDS). Id. It is administered in combination with leucovorin (folinic acid), which provides a source of reduced folate necessary for normal cell function. Leucovorin is readily transported into mammalian cells by an active, carrier-mediated process, and can be assimilated into cellular folate pools following its metabolism. Because the Pneumocystis carinii organism lacks the reduced folate carrier-mediated transport system, leucovorin is prevented from entering the organism. The adjunctive administration of trimetrexate and leucovorin thus protects normal host cells, but not Pneumocystis carinii, from the cytotoxicity of trimetrexate.
Trimetrexate also possesses in vitro and in vivo activity against a range of murine and human tumor cell lines. It shows, for example, in vitro antitumor activity against murine cell lines such as L1210, L5178Y, S-180, W-256, and in vivo utility against murine tumors such as B16 melanoma, colon 26 and 38, L1210 and P388 leukemia and CD8F mammary tumors. Bertino, J. R., et al., Biochem. Pharmacol. 28:1983-1987 (1979); Lin, J.
T., and Bertino, J. R., J. Clin. Oncology 5(12): 2032-2040 (1987); O'Dwyer, P. J., et al., NCI Monographs 5:105-109 (1987). Trimetrexate also exhibits in vitro activity against human tumor cells lines derived from breast, colon, lung, ovary, renal and melanoma cells. Other possible uses for trimetrexate include the treatment of malaria, psoriasis, and rheumatoid arthritis.
The low water solubility (&lt;0.1 mg/ml) and long term instability of trimetrexate free base diminish its usefulness in the treatment of disease, but the severity of these problems can be lessened if trimetrexate salts are instead used. Examples of some trimetrexate salts are disclosed by: Hempel, A., et al., Cancer Biochem. Biophys., 10, 25-30 (1988); U.S. Pat. Nos. 5,716,968 and 5,716,960, both to Kennedy; and U.S. Pat. No. 4,376,858 to Colbry ("Colbry"). Colbry teaches that a preferred salt is trimetrexate glucuronate because of its superior water solubility (&gt;50 mg/ml), stability, and the low toxicity of glucuronic acid. The synthesis of trimetrexate glucuronate is disclosed both by Colbry and by Hicks, J. L., et al., J. Labeled Compounds Radiopharm. 29, 415 (1991).
Although trimetrexate glucuronate oxidizes more slowly than does trimetrexate free 5 base, the solution half-life of trimetrexate glucuronate is reportedly only 51.6.+-.0.8 days at 37.degree. C. Stetson, P. L., et al., J Chromatography 464, 163-171 (1989). There thus remains a need for pharmaceutically acceptable forms of trimetrexate that have formulation advantages such as acceptable solubility and improved stability as solids and/or liquids.